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Ubiquitin Chain Analysis by Parallel Reaction Monitoring
Published on: June 17, 2020
Conformaciones en la ubiquitina desnaturada con urea y en la proteína G a partir de las constantes de acoplamiento
Navratna Vajpai1, Martin Gentner, Jie-Rong Huang
1Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland.
Journal of the American Chemical Society
|February 17, 2010
Resumen
Este estudio introduce nuevos métodos de RMN para medir con precisión las conformaciones de la cadena lateral en proteínas desplegadas, revelando interacciones específicas de la secuencia y mejorando los modelos estructurales.
Área de la Ciencia:
- La biofísica es la biofísica.
- Biología Estructural Biología Estructural
- Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN) Espectroscopia de Resonancia Magnética Nuclear (RMN)
Sus antecedentes:
- La caracterización de las conformaciones de la cadena lateral en proteínas desplegadas es un desafío debido a la escasa dispersión de resonancia en RMN.
- Existe información limitada sobre los detalles dinámicos y estructurales de las cadenas laterales de proteínas en estados desordenados.
Objetivo del estudio:
- Desarrollar y validar esquemas de RMN optimizados para detectar acoplamientos escalares y acoplamientos dipolares residuales (RDC) en proteínas desplegadas.
- Para determinar con precisión las conformaciones de la cadena lateral (ángulo chi) y explorar las interacciones específicas de la secuencia en proteínas desnaturalizadas con urea.
Principales métodos:
- Secuencias de pulso de RMN optimizadas para la detección de acoplamientos escalares (3) J ((HalphaHbeta), (3) J ((NHbeta), (3) J ((C'Hbeta) y (1) D ((CbetaHbeta) RDCs.
- Aplicación a la ubiquitina desnaturada con urea y a la proteína G.
- Interpretación de los acoplamientos utilizando un modelo de rotamer mixto escalonado chi(1).
Principales resultados:
- Detección de hasta seis (3) J-acoplamientos a (1) H ((beta), definiendo chi ((1) ángulos con alta precisión (<2% para la mayoría de los residuos).
- Identificación de desviaciones significativas de los promedios de la bobina, indicando interacciones específicas de secuencia y residuos, particularmente para la serina y la treonina.
- Buena correlación entre los RDC medidos (1) D ((CbetaHbeta) y un modelo de bobina, lo que respalda su validez como primera aproximación.
Conclusiones:
- Los métodos de RMN desarrollados permiten la caracterización rigurosa de las conformaciones de la cadena lateral y de la columna vertebral en proteínas desplegadas.
- Las interacciones específicas de la secuencia influyen significativamente en las conformaciones de la cadena lateral más allá de los modelos de bobina simples.
- Los datos de RDC pueden refinar las descripciones de conformación de la columna vertebral local en estados desplegados.
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